The LC-MS-MS Core will provide analytical support using liquid chromatography-tandem mass spectroscopy (LC-MS-MS) for all projects within this chemoprevention program. LC-MS-MS will be used to characterize active natural products contained in biologically active plant extracts with respect to molecular weight, chromatographic retention time, UV/vis absorbance (with in-line absorbance detection) and structure (based on MS-MS fragmentation patterns). These data will contribute to the structural elucidation of novel active compounds as well as identification of known compounds (a process known as dereplication) in support of Project 2. Because these analyses are fast (typically less than 1 hour each), they may be used to provide feedback to guide the compound scale-up activities. In addition, the purity, molecular weight, and structure of synthetic reference compounds and intermediates prepared will be confirmed by using LC-MS-MS. Whether providing data for the structural elucidating of new active natural products, identification of known natural products, or confirmation of the structures of synthetic compounds, LC-MS-MS will significantly improve the efficiency of several projects. Another function of the LC-MS-MS Core will be to contribute to the bioassay efforts by developing and carrying out screening assays using pulsed ultrafiltration mass spectrometry, which is a screening method invented in this laboratory. Specifically, chemopreventive natural products contained in plant extracts will be identified using pulsed ultrafiltration mass spectrometry based on their binding to cyclooxygenase. Once identified, the affinity constants of these natural products may be determined using pulsed ultrafiltration. Finally, the LC-MS-MS Core will interact with Projects by supporting studies on the mechanism of action of lead chemoprevention agents. For example, an LC-MS-MS assay developed in this laboratory will be used to quantitatively determine the antioxidant activity of lead compounds by measuring their ability to prevent oxidation of specific DNA bases. Since DNA oxidation might result in carcinogenesis or tumor promotion, the prevention of oxidative stress by natural products is a potential mechanism of chemoprevention. In summary, the LC-MS-MS Core will interact will all five Projects within this program by quickly providing essential mass spectroscopic data on active natural products and, as a result, this core facility will significantly enhance the overall efficiency of the program.